Ventricular Septal Defect with Aortic Valve Insufficiency in a New Zealand White Rabbit^S

Introduction

A ventricular septal defect (VSD) occurs if embryonic interventricular communications do not terminate during embryonic and postnatal life.^1–3 This condition is well-known in humans as well as in dogs and cats.^1,4–6 The frequency of isolated VSDs in dogs with congenital heart disease is estimated to be about 7%, whereas in cats, the lesion has been observed in approximately 15% of those with congenital cardiac anomalies.³ Most VSDs are located in the upper part of the ventricular septum and are termed perimembranous (or paramembranous), involving mainly the membranous part and at their periphery, which is the muscular part of the septum as well. Viewed from the left ventricular cavity, perimembraneous defects are referred to as subaortic, located immediately beneath the aortic valve. From the right ventricular side, perimembraneous VSDs are called supracristal if they open distal to the crista supraventricularis ridge just below the pulmonic valve. Infracristal perimembraneous defects open proximal to the crista supraventricularis muscle, close to the tricuspid valve.^2,6,7

A perimembranous VSD might occur together with aortic regurgitation as the VSD is located immediately below the aortic valve, undermining the support for the aortic valve cusps.² This condition occurs in humans and was also described in dogs.^4,7–9

In the rabbit, experimentally induced cardiovascular disorders, such as models of pressure and volume overload ventricular hypertrophy, have been reported.¹⁰ However, descriptions of naturally occurring cardiac diseases in rabbits are rare and most of them are reviews or are case reports.^11–14 Pathologic findings of a rabbit with VSD, Eisenmenger's syndrome, and congenital valvular cysts were reported in a case study.¹⁵ Clinical findings, including phonocardiography, electrocardiography and radiography, as well as necropsy, were published only in one case report by Kanemoto and Chimura.¹⁶ To the best of the authors' knowledge, no description on echocardiographic findings of VSD in rabbits can be found in the literature. The purposes of the present report were to describe the clinical findings and to demonstrate the unique diagnostic capabilities of echocardiography in a case of a New Zealand White rabbit with VSD and aortic regurgitation.

Case Report

A 10 mo old, female New Zealand White rabbit weighing 3.8 kg was obtained for the Small Animal Clinic (SAC) of UVM Hanover from a commercial vendor^a for use in an orthopedic study of tibial magnesium–calcium screws for evaluating screw stability, degradation, and biocompatibility. These experiments were conducted in accordance with the “European Directive for the Protection of Vertebrate Animals Used for Experimental and Other Scientific Purposes (86/609) EU”. The Committee for Experiments on Animals of the University of Veterinary Medicine Hanover approved the experiments. A cardiac murmur was detected during routine clinical examination and further cardiologic examinations were performed to reveal the cardiac diagnosis and to estimate the risk of anesthesia needed for the planned experiment.

Clinical examination demonstrated normal mucosal membranes and a capillary refill time of 2 sec. Body temperature was not measures because stress caused by rectal measurement greatly influenced the temperature and could have serious consequences in rabbits.¹⁷ The respiratory rate was 70 breaths/min (reference values, 30–60 breaths/min) and the heart rate was 190 beats/min (reference values, 180–250 beats/min).¹⁴ Respiratory sounds were slightly increased. A precordial thrill was felt over the ventral third of the left hemithorax. Thoracic percussion detected no abnormalities, such as pleural effusion, pneumothorax, or lung consolidation. Cardiac auscultation revealed a grade 5/6 systolic, widely radiating murmur best heard at the ventral third of the left hemithorax at the fourth intercostal space. The murmur seemed to be extended over systole, partially covering the first part of the second heart sound. A grade 4/6 systolic murmur was detected on the right hemithorax with its point of maximal intensity close to the sternum, at the fourth intercostal place. Strong femoral pulses were palpated on the femoral arteries without signs of arrhythmia or pulse deficit. Routine hematology and serum biochemistry values were within normal reference ranges.¹⁷

Electrocardiography was performed as described previously by others.^11,14,18 Briefly, the rabbit was placed in right lateral recumbency with minimal restraint and without sedation, and standard limb electrodes were used. Electrocardiography demonstrated a normal sinus rhythm with a heart rate of 206 beats/min as well as tall R waves and large T waves in lead II and deep S waves in leads II, III, and aVF. (Figure 1 and Table 1). Standard right lateral and ventrodorsal thoracic radiographs showed generalized cardiomegaly with dorsal displacement of the thoracic trachea. Pulmonary arteries and veins seemed to be somewhat prominent, being suggestive of pulmonary overcirculation.

Table 1 Electrocardiographic Findings in a Rabbit with Ventricular Septal Defect and Aortic Valve Insufficiency

^*From Reusch.¹⁴

^†From Rezakhani and Rezaian.¹⁸

^§Electrocardiography was done in right lateral recumbency with minimal physical restraint, without sedation and after 20 sec of acclimatization. Measurements were performed three times, and the average values are shown.

mV, millivolt; P-wave, the first wave representing the depolarization of the atria on the EKG; PR, the time required for an impulse to travel from the SA node to the ventricle; QRS complex, depolarization of the ventricles; QT, time interval measured from the onset of the Q-wave to the end of the T-wave; Q-wave, the first negative deflection that precedes an R wave; R-wave, the first positive deflection of the QRS complex; S-wave, the first negative deflection that follows an R-wave; SE, standard error; ST-segment, time interval from the end of the QRS interval to the onset of the T-wave; T-wave, the first major deflection following the QRS complex

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M-mode, two-dimensional (2-D), and Doppler echocardiography were done with an ultrasound system and using a 7.0 MHz electronic sector transducer and with simultaneous electrocardiographic recording^b as described for rabbits.^11,19 M-mode echocardiographic measurements were performed according to American Society of Echocardiography (ASE) standards.²⁰ Two-dimensional echocardiographic and spectral Doppler measurements were done as previously described for dogs.^21,22 Results of M-mode, 2-D, and spectral Doppler echocardiographic measurements compared with literary reference values from rabbits are shown in Table 2. M-mode echocardiography revealed moderate left ventricular dilatation as well as normal septal and left ventricular free wall thicknesses. Fractional shortening was normal (37.1%) compared with reference values (Table 2), indirectly reflecting normal contractility of the left ventricle. Two-dimensional 2-D echocardiography demonstrated dilatation of all cardiac chambers. Mild left atrial enlargement was estimated by increased left atrium-to-aortic root ratio of 1.6 compared with reference ranges (Table 2). A VSD of 2 mm in diameter was detected, best seen in the right parasternal four-chamber view, just below the aortic root. Displacement of the aortic root toward the right ventricle and protrusion of the aortic valve into the defect was also observed (see supplementary Video I). The VSD could also be seen in the right parasternal short-axis view, just below the aortic valve (see supplementary Video II) and in the left apical five-chamber (outflow) view (see supplementary Video III). The main pulmonary artery was dilated without morphologic alterations of the valve.

Table 2 M-mode, 2-D, and Spectral Doppler Echocardiographic Findings in a Rabbit with VSD and Aortic Valve Insufficiency

^†From ** Kattinger P et al.¹¹ Based on M-mode measurements of nonsedated pet rabbits aging from 8 mo to 6 yr with a mean age of 3.5 yr (n=13).

^*From * Marini et al..¹⁹ Based on M-mode measurements of 7 mo old, sedated Dutch belted rabbits (n=6).

^§Nonsedated values. Measurements were performed three times and the average values are shown.

Rows 2–9: M-mode echocardiographic measurements; rows 10–12: two-dimensional 2-D echocardiographic measurements; rows 13–21: spectral Doppler echocardiographic measurements (meters per second).

A, late diastolic wave; Ao_D, aortic root diameter in diastole; E, early diastolic wave; EPSS, E point to septal separation; FS, fractional shortening; IVS_ED, interventricular septal thickness at the end of diastole; IVS_ES, interventricular septal thickness at the end of systole; LA_S, left atrial diameter in systole; LVID_ED, left ventricular internal diameter at the end of diastole; LVID_ES, left ventricular internal diameter at the end of systole; LVW_ED, left ventricular posterior wall thickness at the end of diastole; LVW_ES, left ventricular posterior wall thickness at the end of systole; SD, standard deviation; VSD, ventricular septal defect

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Color flow (CF) Doppler echocardiography demonstrated the VSD with left to right shunt, resulting in a disturbed systolic flow best seen in the right parasternal long-axis view (Figure 2, see supplementary Video I). A laminar left-to-right diastolic flow through the defect was also detected in this view and a regurgitant flow from the aorta into the left and right ventricles was seen, indicating moderate aortic valve regurgitation into both ventricles (see supplementary Video I). The shunted blood could also be visualized in the right parasternal short-axis view, at the aortic level (see supplementary Video II) as well as in the left apical five-chamber view (see supplementary Video III). Mild pulmonic valve insufficiency was also demonstrated. The results of the spectral Doppler echocardiographic measurements are shown in Table 2 compared with reference ranges. Peak flow velocity across the shunt measured with continuous Doppler echocardiography revealed a 4.0 m/sec peak flow velocity and the estimated left-to-right ventricular pressure gradient was 64.6 mm Hg (Figure 3). Increased aortic peak flow velocity of 2.0 m/sec was measured and moderate aortic regurgitation with a maximal regurgitant flow velocity of 2.0 m/sec was detected and best interrogated in the left apical five-chamber view (Figure 4). The pulmonary valve peak flow velocity yielded 2.1 m/sec, indicating relative pulmonic stenosis, without valvular alterations. A mild pulmonic insufficiency with a 1.5 m/sec regurgitant flow was also measured. The peak inflow velocities at the mitral and tricuspid valves were somewhat higher in comparison with reported values (Table 2). Based on these echocardiographic findings, a VSD located in the upper part of the septum, immediately beneath the aortic valve and with left-to-right shunting, was diagnosed. Moderate aortic and mild pulmonic insufficiencies were also demonstrated.

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During the 3 wk after the first cardiologic examination, the rabbit showed a good general status without signs of congestive heart failure. Therefore, it was decided to keep the animal within the experimental group of the orthopedic surgery. The rabbit was anesthetized with 15 mg/kg ketamin^c and with 18 mg/kg IM medetomidin^d, then followed after intubation by 2–3 vol% isoflurane^e in an oxygen mixture (0.5 L/min 100% O₂) for the experimental orthopedic surgery and in the same manner for two follow-up microcomputed tomographic scans looking for bone alterations. There were no complications during anesthesia and the animal woke up without any problems. After the second microcomputed tomographic investigation, at the end of the investigation period, inhalation anesthesia was ceased and echocardiography was performed under the same sedation with ketamin and medetomidin. The heart rate slowed down from 240 to 122 beats/min, allowing better visualization of the intracardiac structures with a better time resolution both during 2-D and CF Doppler echocardiography. Basically, the same alterations were seen as in the nonsedated stage, but the VSD and the aortic root as well as aortic valve movements could be better observed (see supplementary Videos I–III). The results of the Doppler examinations were similar as in the nonsedated stage, except that a mild tricuspid regurgitation was also detected by the CF Doppler echocardiography, best depicted in the left apical five-chamber view.

Then, the rabbit was euthanized with 250 mg/kg pentobarbital^f given intracardially, as planned previously for the orthopedic experiment, and bone samples were taken. The thorax was opened from the left side, and the heart and lungs were removed and were fixed in 10% neutrally buffered formalin for later dissection and photography. No pathologic alterations of the other organs were found during gross pathology. Generalized cardiac enlargement was seen with a heart weight of 22 g compared with a healthy rabbit with a similar age and living weight of 3.6 kg and with a postmortem heart weight of 12 g measured after the same fixation method (Figure 5). The left and the right atria were dilated, whereas both ventricles were dilated, showing an increase in thickness with a mild attenuation of the papillary muscles, indicating moderate eccentric hypertrophy and moderate dilatation. These macroscopic alterations were prominent, compared with the healthy control animal shown on Figure 5. The main pulmonary artery was dilated, whereas the pulmonary valves did not show any gross lesions. A VSD of 5×2 mm was observed in the upper, membranous portion of the interventricular septum, as viewed from the left ventricle and just below the noncoronary cusp of the aortic valve, which lacked its structural support at the septal part of the aortic root (Figure 6). The noncoronary and the right coronary cusps of the aortic valves were distorted. Slight rightward displacement of the aortic root was also observed. The defect could also be seen from the side of the right ventricle, just above the right bicuspid valve. The atrioventricular valves were normal. Cardiac histopathology showed left and right ventricular cardiomyocyte degeneration (i.e., swelling and hypereosinophilia of the cytoplasm with a loss of cross striation, and nuclear hyperchromasia), cartilaginous metaplasia of the aorta, and subendocardial fibrosis of the right ventricular flow tract.

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Discussion

Pathophysiology and clinical signs of VSDs mainly depend on the location and size of the defect. Most of them are accompanied with a left-to-right shunt, when blood flows from the left ventricle to the right ventricle, especially during ventricular systole, but also during ventricular diastole, due to ventricular pressure differences.^2,3,6 Only large VSDs coincide with a right-to-left or bidirectional shunt and with a high pulmonary resistance, either from birth or developing later from a left-to-right shunt, resulting in Eisenmenger's syndrome. The size of the VSDs are classified as small, medium, or large shunts.^2,6 The most typical small- to medium-sized VSDs are located high in the membraneous septum, allowing the shunted blood to be ejected from the right ventricle directly into the right ventricular outflow tract and out to the main pulmonary artery. Thus, the right ventricle is more or less spared from the higher workload during systole and only the left ventricle has ventricular volume overload, coping with the extra amount of blood arriving from the overcirculated pulmonary vasculature toward the left atrium. Small defects are commonly termed restrictive VSDs, meaning that the size of the defect restricts the higher left ventricular pressure from being transmitted to the right ventricle, and it allows only a relatively limited amount of blood to flow across the shunt. The high pressure difference results in a high velocity left-to-right disturbed flow during systole, but low velocity shunting can also occur during diastole.^2,3 Moderate size to large left-to-right shunts result in both left and right ventricular volume overload hypertrophy as well as overcirculation of the pulmonary vasculature and pulmonary hypertension. Large VSDs that have the same or greater surface area as the aortic valve region provide only little resistance against the flow of the shunting blood through the defect.

This rabbit was diagnosed with a congenital heart defect, causing a heart murmur and cardiac enlargement, based on the history, physical findings, electrocardiography, and radiography. The rabbit did not show clinical signs of congestive heart failure, despite its cardiac alterations. These findings are similar to dogs and cats with small- to moderate-sized VSDs.^3,9 This might also be because laboratory and pet rabbits are usually not prone to exercise.

Kanemoto and Chimura reported a rabbit with VSD and a pansystolic murmur recorded by phonocardiography.¹⁶ In the authors' case, the murmur on the left side was different compared with that on the right side and seemed to have both systolic and diastolic components during auscultation. A similar, systolic–diastolic murmur was recorded with phonocardiography above the left side of the thorax by Kettner et al. in a dog with VSD and with quadricuspid aortic valve, causing aortic regurgitation.²³

In dogs and cats, electrocardiographic and radiographic findings highly depend on the characteristics, such as the size and location of the VSD as well as pulmonary vascular resistance.^3,6,9 The electrocardiogram of this case differed from those of normal rabbits as it showed tall R waves and large T waves in lead II as well as deep S waves in leads II, III, and aVF. Forms and sizes of the QRS complexes are greatly variable in rabbits.¹⁸ However, the tall R waves seen in this rabbit might be considered as signs of left ventricular enlargement, which was supported by the echocardiographic and postmortem findings. Thoracic radiographic findings were similar to the canine cases with moderate-sized VSDs combined with aortic insufficiency.^7–9 In an earlier report, a VSD in a rabbit was accompanied by electrocardiographically suggested left ventricular and radiographically detected biventricular enlargement as well as with dilated pulmonary vessels.¹⁶

Searching through the scientific literature, the authors did not find any publications on echocardiographic findings of congenital heart defects in rabbits. Echocardiography provided a correct in vivo diagnosis of a VSD in this case, similarly to previous reports on dogs.^7–9 The size of a VSD can be determined by 2-D echocardiography, whereas Doppler echocardiography is a valuable, noninvasive modality to estimate shunt magnitude.^2,3 With the help of continuous wave Doppler echocardiographic measurements, VSDs can be categorized by applying the modified Bernoulli equation for calculating the systolic pressure gradient between the left and right ventricles from the maximal flow velocity across the shunt. The jet velocity for a completely restrictive defect with a high-pressure gradient was estimated by Kittleson as between 4.6 and 5.8 m/sec.² Oyama et al. found the jet velocity through restrictive defects as equal or greater than 4.5 m/sec.⁶ Tidholm reported peak shunt flow velocity between 4.0 and 5.0 m/sec in 13 canine cases with VSD, and one of them with aortic regurgitation, without mentioning shunt size or flow velocity categories.⁵ In the authors' case, the highest shunt velocity yielded 4.0 m/sec with 64.6 mm Hg (both in nonsedated and sedated stage), which might represent a moderate left-to-right shunt compared with the literary data for canine VSD. This shunt magnitude seemed to provide a partial explanation of the other cardiologic consequences, such as biventricular volume overload, dilatation of the pulmonary artery, and pulmonary overcirculation similar to reported canine cases. Shunt fraction Qp:Qs calculation could have provided further information in this respect but was not performed.²⁴ VSDs located immediately below the aortic valve can undermine the structural support of the aortic valve apparatus and the aortic sinuses, resulting in aortic regurgitation.^2,4,6–8 The echocardiographically revealed aortic insufficiency might have contributed to the pathophysiology of this case.^7–9 The authors did not find any gross pathology alteration (e.g., subvalvular, valvular, or supravalvular stenosis of the pulmonary artery), which might have provided an explanation of the enlargement of the pulmonary artery. It can only be speculated that the increased pulmonary flow and mild regurgitation could be partially due to the increased blood flow in the right ventricular tract originating from the shunt and from aortic insufficiency partially regurgitating into the right ventricle. Left-to-right shunts with consequent elevation of the right ventricular pressure might result in a pressure-related subendocardial fibrosis of the truncus pulmonalis.²⁵ This histopathologic alteration was also seen in the authors' case.

The authors measured the size of the defect by 2-D echocardiography as 2×2 mm in diameter. The real size of the VSD proved to be of 5×2 mm during gross pathology, which could be the different postmortem morphologic stage of the heart, but was otherwise because 2-D echocardigraphy often underestimates the size of a small VSD due to its limitations in lateral resolution.⁹

Displacement of the aortic root and prolapse of the aortic valve were also seen during 2-D echocardiography, similar to previous reports on humans and dogs.^4,8,9,23 Tetralogy of Fallot with overriding aorta and pulmonic stenosis should be a differential aspect of a VSD with secondary aortic regurgitation. In the authors' case, tetralogy of Fallot was excluded based on the echocardiographic findings and also by pathology.

As to the authors' observations, sedation might be suggested to avoid unnecessary movements of the patient and to slow down the heart rate for better visualization of the cardiac structures and events. As rabbits with cardiac diseases might have risks with anesthesia, close monitoring and proper anesthesia is needed during this process. In the rabbit, the size of the VSD was reported to be 1.5–3 mm based on pathology findings of two case reports in comparison with the authors' case of 2×5 mm.^15,16 In a postmortem study, the heart weights of 348 normal rabbits varied between 3.2 and 10.2 g.²⁶ The heart weight of the authors' case measured at necropsy yielded 22 g, which was well over these values and was due to compensatory ventricular hypertrophy. Also, the relative heart weight of this rabbit was 0.58%, whereas this proportion varied between 0.22% and 0.27% in an experimental study performed on New Zealand White rabbits.²⁷

In addition to the few reported natural cases of VSD in rabbits, it is known that nonsteroid anti-inflammatory drugs can cause VSD as an embryonic malformation if the pregnant rabbit is treated with some of these drugs (e.g., aspirin.²⁸ This was not the situation regarding the authors' case and the etiology of the defect could not be cleared, as none of the littermates or the mother of the patient had similar problems, to the authors' knowledge.

Conclusion

In the present case, a congenital cardiac defect was suspected in a rabbit with cardiac murmur and without other clinical signs. Echocardiographic findings of congenital heart defects have not been reported in rabbits. In this case, a VSD with aortic insufficiency was diagnosed by 2-D and Doppler echocardiography. The latter modality was also useful in determining the severity of the VSD, similar to earlier canine reports.